Detection of only specific cells from a sample containing various types of cells is needed in a wide range of field. Particularly, in the biotechnological research and development and in the clinical practice associated with medicine and pharmacy, checking large numbers of cells for the presence or absence of specific cells (for example, cells containing or expressing a specific substance, cells infected with pathogens, and cancer cells) is very important for the progress of experiment and for disease diagnoses.
However, detection of specific cells from a sample containing large numbers of cells has been difficult, particularly when the percentage of the specific cells is low.
For example, in malaria parasite infection caused by the infection of red blood cells with malaria parasites, the following methods are available for the detection and diagnosis of the infection.
For example, in one method, blood cells are smeared on a glass slide, dried, and observed under a microscope after Giemsa staining to find the presence or absence of a malaria parasite infection or the extent of the infection. This method is advantageous, because it requires only the staining procedure for microscope observation, and is therefore easy to perform. The method, however, requires time and sophisticated observation techniques for the detection and diagnosis, and the detection sensitivity is very low. For example, the method takes as long as 40 minutes, and the detection sensitivity (detection limit) is only just high enough to detect infected cells accounting for at least about 0.01% of the whole blood cells.
In another method, the presence or absence of a malaria parasite infection or the extent of the infection is tested or diagnosed using immunochromatography. The method is widely known, and test kits are commercially available to enable easy testing. Such test kits offer even simpler procedures for observing the presence or absence of an infection or the extent of the infection. The immunochromatography takes only about 20 minutes; however, the detection sensitivity is only at the levels of microscope observation. Further, since the immunochromatography is prone to showing false positive results, it is common practice to combine the method with microscope observation.
Another method uses PCR to test and diagnose the presence or absence of a malaria parasite infection or the level of the infection. Unlike the microscope observation, the method offers high detection sensitivity. However, the PCR requires about 5 hours to produce results, and the procedure is complex and requires high skills. Further, depending on sample conditions and primers, setting reaction conditions is difficult, and detection may not be possible. The use of PCR alone is thus not suited for the diagnosis of infections.
Thus, in malaria parasite infection taken as an example above, the conventional detection and diagnosis methods have a number of drawbacks, including long hours, high skills, and poor detection sensitivity. Accordingly, there is a need for a novel infection detection method that is rapid, easy, and highly sensitive.
To date, various types of microarray chips have been reported, and used for various purposes, including cell detection. For example, Patent Literature 1 reports a system in which microchambers are integrated and connected to a heater and other parts of the system to enable high-throughput biochemical reactions of a biological sample. However, the system is not sensitive enough to enable detection at a single cell level, and high-sensitive detection of small numbers of target cells contained in large numbers of cells is difficult.
Patent Literature 2 reports a technique for observing the effect of a substance on cells. This is achieved by patterning a polymer on a substrate, and allowing cells to act on a substance of interest immobilized on the polymer. However, it is difficult with this technique to immobilize a certain number of cells on each pattern, and to detect small numbers of target cells in larger numbers of cells with high sensitivity.
Patent Literature 3 reports an array chip that includes large numbers of integrated microwells designed to contain a single lymphocyte per microwell. However, since only a single cell can be introduced to each microwell, screening of cells that outnumbers the microwells integrated on the array chip is difficult. Accordingly, it is difficult with this array chip to perform high-sensitive detection of small number of target cells contained in large numbers of cells (1 million to 10 million cells). It is also considered very difficult to culture the introduced cells or to perform PCR for these cells with the microwells of this array chip.
As described above, high-sensitive detection of small numbers of target cells (specific cells) contained in large number of cells is difficult to achieve with the conventional methods that use the microarray chips.
Under these circumstances, there is a need for a novel detection means that enables fast, easy, and high-sensitive detection of a variety of target cells (specific cells), including, for example, cells infected with various infections such as malaria parasite infection. There is also a need for a novel detection means that can easily be used, for example, at the bedside. However, no such detection means have been reported.